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The isolation of circulating tumoral DNA (ctDNA) present in the bloodstream brings about the opportunity to detect genomic aberrations from the tumor of origin. However, the low amounts of ctDNA present in liquid biopsy samples makes the development of highly sensitive techniques necessary to detect targetable mutations for the diagnosis, prognosis, and monitoring of cancer patients. Here, we employ standard genomic DNA (gDNA) and eight liquid biopsy samples from different cancer patients to examine the newly described CRISPR-Cas13a-based technology in the detection of the BRAF p.V600E actionable point mutation and appraise its diagnostic capacity with two PCR-based techniques: quantitative Real-Time PCR (qPCR) and droplet digital PCR (ddPCR). Regardless of its lower specificity compared to the qPCR and ddPCR techniques, the CRISPR-Cas13a-guided complex was able to detect inputs as low as 10 pM. Even though the PCR-based techniques have similar target limits of detection (LoDs), only the ddPCR achieved a 0.1% variant allele frequency (VAF) detection with elevated reproducibility, thus standing out as the most powerful and suitable tool for clinical diagnosis purposes. Our results also demonstrate how the CRISPR-Cas13a can detect low amounts of the target of interest, but its base-pair specificity failed in the detection of actionable point mutations at a low VAF; therefore, the ddPCR is still the most powerful and suitable technique for these purposes. [ABSTRACT FROM AUTHOR]

Simple Summary: The prognosis of patients with stage II and stage III colon cancer is heterogeneous. Clinical and pathological characteristics may help to further refine the recurrence risk. We built a prognostic score and categorized patients into two risk groups in a training and validation cohort. We assigned two points to T4 and one point to N2 and high tumor budding based on the multivariate cox regression analysis for time to recurrence (TTR) in the training cohort. Forty-five percent of the patients were assigned to the low-risk group and compared to the high-risk group, had a significantly longer TTR. These results were confirmed in the validation cohort. Background: The prognosis of patients with stage II and stage III colon cancer is heterogeneous. Clinical and pathological characteristics, such as tumor budding, may help to further refine the recurrence risk. Methods: We included all the patients with localized colon cancer at Hospital Universitario La Paz from October 2016 to October 2021. We built a prognostic score for recurrence in the training cohort based on multivariate cox regression analysis and categorized the patients into two risk groups. Results: A total of 440 patients were included in the training cohort. After a median follow-up of 45 months, 81 (18%) patients had a first tumor recurrence. T4, N2, and high tumor budding remained with a p value <0.05 at the last step of the multivariate cox regression model for time to recurrence (TTR). We assigned 2 points to T4 and 1 point to N2 and high tumor budding. Forty-five percent of the patients were assigned to the low-risk group (score = 0). Compared to the high-risk group (score 1–4), patients in the low-risk group had a significantly longer TTR (hazard ratio for disease recurrence of 0.14 (95%CI: 0.00 to 0.90; p < 0.045)). The results were confirmed in the validation cohort. Conclusions: In our study, we built a simple score to predict tumor recurrence based on T4, N2, and high tumor budding. Patients in the low-risk group, that comprised 44% of the cohort, had an excellent prognosis. [ABSTRACT FROM AUTHOR]

Simple Summary: The survival of colorectal cancer patients largely relies on the stage at diagnosis. The identification of early and non-invasive biomarkers to be used in screening programs for the diagnosis of the disease at the premalignant stage is mandatory. The aim of this study is to validate in plasma-derived extracellular vesicles secreted by malignant cells the diagnostic potential of well-known tumor-associated genes, ΔNp73, TAp73, and Δ133p53, in healthy subjects (n = 29), individuals with premalignant lesions (n = 49), and colorectal cancer patients (n = 42). Our data support ΔNp73 levels contained in extracellular vesicles as such a non-invasive and premature biomarker for the early diagnosis of colorectal cancer. The early diagnosis of colorectal cancer is a key factor in the overall survival of the patients. The actual screening programs include different approaches with significant limitations such as unspecificity, high invasiveness, and detection at late stages of the disease. The specific content of extracellular vesicles derived from malignant cells may represent a non-invasive technique for the early detection of colorectal cancer. Here, we studied the mRNA levels of ΔNp73, TAp73, and Δ133p53 in plasma-derived extracellular vesicles from healthy subjects (n = 29), individuals with premalignant lesions (n = 49), and colorectal cancer patients (n = 42). Extracellular vesicles' ΔNp73 levels were already significantly high in subjects with premalignant lesions. Δ133p53 levels were statistically increased in colorectal cancer patients compared to the other two groups and were associated with patients' survival. Remarkably, TAp73 mRNA was not detected in any of the individuals. The evaluation of ΔNp73, Δ133p53 and CEA sensitivity, specificity and AUC values supports ΔNp73 as a better early diagnosis biomarker and CEA as the best to identify advanced stages. Thus, low levels of CEA and a high content of ΔNp73 may identify in screening programs those individuals at higher risk of presenting a premalignant lesion. In addition, Δ133p53 emerges as a potential prognosis biomarker in colorectal cancer. [ABSTRACT FROM AUTHOR]